Bacterial resistance to mercuric ions and orgnaomercurials is due to the production of two different enzymes, the mercuric reductase which reduces Hg2+ to Hg0, and the organomercurial hydrolase which breaks the Hg-C bond. The Hg0 produced by the presence of these enzymes volatilizes, thus resulting in detoxification of the micro-environment. From preliminary evidence, a similar detoxification mechanism is hypothesized to be present in marmalian systems. The objective of the proposed research is to isolate the detoxifying enzymes and characterize the molecular mechanism by which they transform mercuric compounds. The characterization of these enzyme catalyzed inorganic reactions will be of fundamental importance to biochemistry and biological metal detoxification. The isolation will be based on a published procedure, incorporating affinity chromatography. Investigations will be made to determine the number and nature of electron accepting groups associated with the purified enzyme. The molecular mechanisms of the detoxifying enzymes will be studied in detail using various kinetic techniques. Steady-state kinetic analysis will allow the quantitation of binding site number and affinities, as well as the detection of covalent intermediates. Pre-steady-state studies may indicate transient catalytic intermediates not easily observed. Anaerobic titrations with various electron donors and acceptors will give information as to the mode of electron transfer. The synthesis of transition-state analogues and suicide substrates based on the kinetic analysis above will also be a part of the proposed study. The localization and isolation of similar mercury detoxifying enzymes in the rat will provide substantial evidence for the existence of this detoxification mechanism in man.